Pneumocystis has risen to the forefront of lethal, opportunistic lung infections as a result of its close association with the HIV epidemic, yet this atypical fungal pathogen is becoming increasingly associated with pharmacologic-driven immunosuppression as well as patients with chronic lung diseases such as COPD. We have recently investigated the role of transcription factor STAT4 in lung immunity to P. murina. We have discovered that Stat4-/- mice on the C57BL/6 background are resistant to P. murina infection while Stat4-/- mice on the BALB/c background are susceptible (Myers et al. submitted, PLoS Pathogens). To our surprise, although having lower Th1 (IFN-y) responses, both BL/6 Stat4-/- and Balb/c Stat4-/- mice unexpectedly demonstrated significantly impaired CD4+ Th2 (IL-4, IL-5 and IL-13) responses in the lung. In spite of this, BL/6 Stat4-/-, but not Balb/c Stat4/-, mice maintained an enhanced M2 macrophage signature (higher Retnla/FIZZ-1 mRNA expression and CCL17/TARC protein levels). In contrast to lung CD4+ T cells, BL/6 Stat4-/- mice demonstrated enhanced CD4+ Th2 responses in the draining lymph nodes and enhanced P. murina-specific IgG1, IgG2b and IgG2c levels in sera, neither of which were observed in BALB/c Stat4-/- mice. Together, these results suggest that protective immunity to P. murina when lung CD4+ T cell responses are impaired involves systemic CD4+ Th2 responses driving optimal B cell responses (Ab production) and a local type 2 response (IL-4/IL-13) in the lungs, which may not be CD4+ T cell in origin, driving M2 macrophage development. This is further supported by new preliminary data showing that compared to non-colonized HIV- infected individuals, Pneumocystis-colonized HIV-infected individuals have significantly lower plasma levels of the Th2 cytokines IL-4, IL-5 and IL-13, but no difference in the levels of IFN-y, IL-17A or IL-10. In preliminary studies, we show that CD4(-)/CD49b(-)/SiglecF(-)/ICOS(+) innate helper type 2 cells were significantly increased in lung tissue of C57BL/6 mice at 2 weeks post-P. murina challenge, suggesting a potential role for this cell population in generating type 2 responses in the lung during P. murina infection. In other studies, employing CD19-DTR mice, which are CD19-Cre mice crossed with Cre-inducible diphtheria toxin receptor (DTR) transgenic mice, we found that CD4+ Th1 and Th2 responses were attenuated in the lymph nodes of mice when B cells were depleted prior to P. murina infection, yet B cell depletion resulted in enhanced CD4+ Th2 and Th17 responses in the lung. Collectively, our data leads us to hypothesize that the generation of type 2 and Th2 responses mediate protective lung immunity against Pneumocystis. To address this hypothesis, we have proposed the following two independent, interrelated Aims: (1) to define the development of local and systemic type 2/Th2 responses during P. murina lung infection and (2) define the role of B cells in the development of local and systemic CD4+ T cell responses during P. murina lung infection.